Inhibition of mTORC1 Kinase Activates Smads 1 and 5 but Not Smad8 in Human Prostate Cancer Cells, Mediating Cytostatic Response to Rapamycin

Although hyper-activated mTOR is well recognized as being pivotal to prostate cancer growth and progression, the underlying mechanisms by which it promotes such responses remain incompletely understood. Here we show that rapamycin activates Smads 1 and 5 in human prostate cancer cells and tissues through blocking mTORC1 kinase. ShRNA-based gene silencing and gene overexpression approaches reveal that Smads 1 and 5 mediate while Smad8 represses rapamycin-induced cell death and expression of the BMP transcriptional target Id1 in human prostate cancer cell lines. Moreover, such phospho-Smad1/5-mediated rapamycin responses were blocked by LDN-193189 (a BMPRI kinase inhibitor) or Noggin (a BMP antagonist) in LNCaP prostate cancer cells. Likewise, the mTOR kinase inhibitors Ku-0063794 and WYE-354 each enhanced phosphorylation of Smad1/5. Intriguingly, silencing Raptor alone enhanced while silencing Rictor repressed the phosphorylation of Smad1/5, indicating that mTORC1 represses while mTORC2 activates BMP signaling. Immunohistochemical analysis showed increased levels of phospho-Smad1/5 concomitant with suppression of phospho-S6 and Survivin levels in PC3 human prostate cancer xenografts in athymic mice administered rapamycin (i.p., 5 mg/kg/day, 2 to 6 days). Moreover, we show that compared to prostate tumor tissue from untreated patients, levels of phospho-Smad1/5 were significantly elevated in the prostate tumor tissue of high-risk prostate cancer patients who received 8 weeks of the rapalog everolimus as part of a neoadjuvant clinical trial prior to undergoing local definitive therapy by radical prostatectomy. Taken together, our data implicate Smads 1, 5 and 8 as potential prognostic markers and therapeutic targets for mTOR inhibition therapy of prostate cancer.